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PNA probes have no direct interaction with DNA polymerase. However, PNAs can terminate oligonucleotide primer elongation by binding to the template or by competing with the primers. This PNA-directed PCR clamping strategy uses PNAs to inhibit amplification of a specific target by direct competition between a PNA targeted against one of the PCR primer sites and the conventional PCR primer. This PNA–DNA complex, formed at one of the primer sites, effectively blocks PCR product formation. The procedure is so powerful that it can detect single-base-pair gene variants for mutation screening and gene isolation. The peptide nucleic acid (PNA) blockers can be used to block amplification from the internal transcribed spacer. For example, PCR can include peptide nucleic acid (PNA) blockers targeting plant plastid and mitochondrial rRNA genes for 16S rRNA gene amplification (GGCAAGTCTTCTTCGGA and GGCTCAACCCTGGACAG) and PNA targeting plant nuclear rRNA genes for the ITS2 region (CGAGGGCACGTCTGCCTGG) to reduce amplification of plant material. |
